Acute kidney injury (AKI) is an enormous medical problem, with a very high incidence and mortality rate in hospitalized patients, especially in the ICU. Treatment consists solely of supportive care and, in extreme cases, renal replacement. A major factor in our failure to find effective therapy is that our understanding of how the kidney recovers from AKI is very limited and so we have been handicapped in identification of possible approaches and drugs for therapy. We have developed an innovative new system to study the recovery from AKI, which will allow us to better analyze the molecular and cellular basis of this process from AKI. This will in turn allow us to identify, test and refine candidate therapies. A major target in AKI is the epithelium, especially proximal tubule cells. Severe insults produce dead cells, which are extruded into the tubular lumen. The focus of this grant is to understand how the tubule is subsequently repaired, with the long term goal of improving repair. We will test the hypothesis that phosphatidylinositols 3,4-bisphosphate (PIP2) and 3,4,5-trisphosphate (PIP3) play specific roles in wound healing. We will use live cell imaging to test the prediction that PIP3 controls both formation of the leading edge during cell spreading, as well as cell height during cell spreading and repolarization at the end of wound healing. We will also test the prediction that PIP2 controls the size of the apical plasma membrane and thereby influences cell shape and spreading. We will confirm this in an in vivo model of AKI. We predict that different isoforms of the enzyme that synthesizes PIP3 may have distinct functions, such as in controlling cell height or formation of the leading edge. When some cells in the tubule die, the surviving cells partially depolarize, dedifferentiate, migrate and proliferate to cover the denuded areas. Later, as the surviving cells make contact with their new neighbors, the cells stop migration and proliferation in the process of contact inhibition. We will test the roles of two signaling pathways, Ras-Raf-MEK-ERK cascade and the PAK-PIX complex, in control of differentiation, migration and contact inhibition. PUBLIC HEALTH RELEVANCE: Acute kidney injury is an enormous medical problem, for which there is no therapy. The absence of therapy is a result of our lack of understanding of how the kidney recovers from injury. We have developed an innovative cell culture model system to study the cellular and molecular mechanisms of recovery from acute kidney injury.